PrestoBlue™ Cell Viability Reagent
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What is PrestoBlue™ Reagent?
PrestoBlue™ reagent is a resazurin-based solution that functions as a cell viability indicator by using the reducing power of living cells to quantitatively measure the proliferation of cells. The PrestoBlue™ reagent contains a cell-permeant compound that is blue in color and virtually nonfluorescent. When added to cells, the PrestoBlue™ reagent is modified by the reducing environment of the viable cell, turns red in color and becomes highly fluorescent. This change can be detected using fluorescence or absorbance measurements.
Assay Workflow Complete in 10 Minutes
A 96-well plate containing the cells and the compounds to be tested is prepared. The PrestoBlue™ reagent is added directly to cells and incubated at 37°C for 10 minutes. The plate is then transferred to a fluorescence reader to measure signal. Results are evaluated by plotting signal versus compound concentration (Figure 1). In addition to this protocol format, the signal can alternatively be detected by absorbance or the assay can be run at room temperature. (View product protocol for full details on these assay variations.)
Figure 1. PrestoBlue™ cell viability reagent protocol.
Reliable, High Quality Results
Figure 2. Excellent data obtained in just 10 minutes. Jurkat cells were plated in cell culture medium in a 384-well plate in quadruplicate starting at 100,000 cells/well with a two-fold serial dilution. Cells were incubated with PrestoBlue™ reagent for 10 minutes prior to fluorescence measurements.
Sensitive Detection of Cell Viability
Figure 3. Extremely sensitive detection. Jurkat cells were plated in cell culture medium in a 384-well plate in quadruplicate starting at 100,000 cells/well with a two-fold serial dilution. Fluorescence measurements were taken following 10 minutes of incubation and following 16 hours of incubation with PrestoBlue™ reagent.
Assay Workflow Complete in 10 Minutes
A 96-well plate containing the cells and the compounds to be tested is prepared. The PrestoBlue™ reagent is added directly to cells and incubated at 37°C for 10 minutes. The plate is then transferred to a fluorescence reader to measure signal. Results are evaluated by plotting signal versus compound concentration (Figure 1). In addition to this protocol format, the signal can alternatively be detected by absorbance or the assay can be run at room temperature. (View product protocol for full details on these assay variations.)
Figure 1. PrestoBlue™ cell viability reagent protocol.
Reliable, High Quality Results
) | Data obtained using PrestoBlue™ reagent shows excellent linearity (R2 value of 0.9948) for cells plated at densities between 100 and 100,000 cells/well. The signal for 100 wells was greater than that from zero cells +/- 3 standard deviations, and the Z’ values were above 0.5 for cell densities of 391 cells/well or greater (Figure 2). |
Sensitive Detection of Cell Viability
) | At 10 minutes of incubation, PrestoBlue™ reagent can detect as few as 98 cells/well, and the signal remains linear over the a broad dilution range. Following a 16-hour incubation with PrestoBlue™ reagent, as few as 12 cells/well exhibited a signal that was greater than that from zero cells +/- 3 standard deviations. Z’-factor values were above 0.5 for cell densities of 24 cells/well or greater (Figure 3). Note that at low cell densities, the signal-to-background ratio increases after 16 hours’ incubation. |
PrestoBlue™ Reagent for Primary and Stem Cells
) | PrestoBlue™ Reagent is Compatible With Primary Cells Cells remain viable so you can perform downstream functional assays with your cells after you have assessed their viability status. PrestoBlue™ reagent is the ideal option when using cells available in a limited supply such as primary cells (Figure 4). |
Figure 4. PrestoBlue™ reagent enables analysis of primary cells. PrestoBlue™ reagent has been tested using human aortic smooth muscle cells.
PrestoBlue™ Comparison to Other Resazurin-Based Viability Assays
) | PrestoBlue™ Reagent Viability Assays Are Significantly Faster Than Rezasurin-based Viability Assays, Without Any Loss of Sensitivity PrestoBlue™ reagent only requires a 10-minute incubation period, whereas alamarBlue® and CellTiter-Blue® recommend a 1 to 4 hours incubation. Figure 5 demonstrates the difference in fluorescence signal obtained from PrestoBlue™ reagent and CellTiter-Blue® reagent following a 10-minute incubation. The dynamic range is significantly greater at the early time points for PrestoBlue™ reagent. |
Figure 5. Achieve assay results with PrestoBlue™ reagent in a fraction of the time. CHO-K1 cells were plated in cell culture medium in a 384-well plate in quadruplicate starting at 10,000 cells/well with a two-fold serial dilution. Cells were incubated with PrestoBlue™ reagent, alamarBlue® reagent, or CellTiter-Blue® reagent for 10 minutes or 4 hours. Fluorescence measurements obtained with PrestoBlue™ reagent in 10 minutes take 4 hours with alamarBlue® and CellTiter-Blue® reagents.
PrestoBlue™ Reagent Compared to Tetrazolium Salt-Based Viability Assays
The PrestoBlue™ Reagent Has Many Advantages Over Tetrazolium Salts Like MTT.
- Faster time-to-results and fewer protocol steps (Table 1)
- Ability to maintain your cells in a living state for downstream assays
- A safer reagent that minimizes toxic waste considerations
- A ready-to-use aqueous solution
Shorter Assay Times
Table 1. Reduced time-to-results with PrestoBlue™ reagent. With PrestoBlue™ reagent, results can be obtained in a minimum of 10 minutes as compared to a minimum of 1 hour for XTT or 2 hours for MTT.
| MTT | XTT | PrestoBlue™ | |
|---|---|---|---|
| Minimum incubation time | 1 hr | 1 hr | 10 min |
| Solubilization time | 1 hr | N/A | N/A |
| Total time | 2 hrs | 1 hr | 10 min |
Table 1. Reduced time-to-results with PrestoBlue™ reagent. With PrestoBlue™ reagent, results can be obtained in a minimum of 10 minutes as compared to a minimum of 1 hour for XTT or 2 hours for MTT.
PrestoBlue™ Comparison to CellTiter® Reagent
) | Continuous Live-cell Monitoring and Assessment After Measuring Viability Comparable results obtained with PrestoBlue™ reagent or CellTiter-Glo® reagent in 10 minutes. PrestoBlue™ reagent does not require cell lysis like the CellTiter-Glo® reagent does. This allows you to leave PrestoBlue™ reagent with the cell samples and read the signal up to 24 hours later. With PrestoBlue™ reagent, cells remain viable so you can perform downstream functional assays with your cells after you have assessed their viability status. If you are using cells that are in limited supply such as primary cells, PrestoBlue™ reagent is an excellent choice. |
Figure 6. Comparable results obtained with PrestoBlue™ reagent or CellTiter-Glo® reagent in 10 minutes. U-2 OS cells were plated in a 384-well plate at 2,000 cells/well. Cells were then exposed to various concentrations of Etoposide, Doxorubicin, Chetomin, or Staurosporine for 72 hours. Subsequently, cells were incubated for 10 minutes with PrestoBlue™ reagent (left panel) or CellTiter-Glo® reagent (right panel) prior to fluorescence (PrestoBlue™ reagent) or luminescence (CellTiter-Glo® reagent) measurements. Comparable EC50 values were obtained for each compound.
For Research Use Only. Not intended for any animal or human therapeutic or diagnostic use.
CellTiter®-Glo and CellTiter®-Blue are registered trademarks of Promega Corporation.
alamarBlue® is a registered trademark of TREK Diagnostic Systems.
CellTiter®-Glo and CellTiter®-Blue are registered trademarks of Promega Corporation.
alamarBlue® is a registered trademark of TREK Diagnostic Systems.
HCEC
- Learn more about Human Corneal Epithelial Cells (HCEC)
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